Warm temperature-sensitive transient receptor potential vanilloid 4 (TRPV4) plays an essential role in thermal hyperalgesia

Animals sense various ranges of temperatures by cutaneous thermal stimuli. Transient receptor potential vanilloid 4 (TRPV4) is a cation channel activated at a warm temperature (over 30 degrees C) in exogenously expressed cells. We found in the present study that TRPV4 is essential in thermal hyperalgesia at a warm temperature in vivo. TRPV4-/- and TRPV4+/+ mice exhibited the same latency of escape from 35-50 degrees C hotplates. Neuronal activity in the femoral nerve, however, revealed that the number and activity level of neurons decreased in response to a warm temperature in TRPV4-/- mice. TRPV4-/- mice displayed a significantly longer latency to escape from the plates at 35- 45 degrees C when hyperalgesia was induced by carrageenan without changes in foot volumes. TRPV4 therefore determines the sensitivity rather than the threshold of painful heat detection and plays an essential role in thermal hyperalgesia.     

Methane and hydrogen production in a termite-symbiont system

Methane and hydrogen emission rates and the ¹³C of CH₄ were observed for various termites in Australia, Thailand and Japan. Combined with the already reported emission rates of CH₄ in the literature, the phylogenetic trend was examined. Emission rates of the observed termites were categorized into five groups: group I with high CH₄ and low H₂ emission rates with a CH₄/H₂ ratio of typically 10/1; group II with high CH₄ and high H₂ emissions with a CH₄/H₂ ratio of 4/1–1/2; group III with low emission rates of CH₄ and H₂; group IV with high H₂ and insignificant CH₄ emissions; and group V with insignificant emissions for both CH₄ and H₂. In lower termites, there are both colonies infected and uninfected with methanogens even in the same species, and no specific trend in CH₄ and H₂ emissions was observed within a genus. Whether protozoa in the hindgut of termites are infected with methanogens or not and the differences in species compositions of protozoa are possibly responsible for the inter-colonial variations. The proportions of infected colonies were possibly small for the family Kalotermitidae (dry wood feeders), and relatively large for families of wet or damp wood feeders. The hydrogen emission rate possibly depends on the locality of methanogens: namely, whether they are intracellular symbionts of protozoa or whether they are attached to the hindgut wall. Emission rates of higher termites were classified into groups according to genera and the diet. Most species of soil or wood/soil interface feeders classified into group I, while the soil feeders Dicuspiditermes in Thailand and Amitermes in Australia were classified into groups with high H₂ emission rates. Typical wood-feeding termites and fungus-growing termites were classified into group III. The results indicate that higher termites tend to increase the CH₄ emission rate during dietary evolution from wood- to soil-feeding, and two types of the system with different efficiencies of interspecies transfer of H₂ have been formed. The ¹³C of CH₄ was discernible with a difference in the decomposition process in the termite–symbiont system among lower termites, fungus-growing termites and other higher termites.     

Fibrillarin, a nucleolar protein, is required for normal nuclear morphology and cellular growth in HeLa cells

Fibrillarin is a key small nucleolar protein in eukaryotes, which has an important role in pre-rRNA processing during ribosomal biogenesis. Though several functions of fibrillarin are known, its function during the cell cycle is still unknown. In this study, we confirmed the dynamic localization of fibrillarin during the cell cycle of HeLa cells and also performed functional studies by using a combination of immunofluorescence microscopy and RNAi technique. We observed that depletion of fibrillarin has almost no effect on the nucleolar structure. However, fibrillarin-depleted cells showed abnormal nuclear morphology. Moreover, fibrillarin depletion resulted in the reduction of the cellular growth and modest accumulation of cells with 4n DNA content. Our data suggest that fibrillarin would play a critical role in the maintenance of nuclear shape and cellular growth.     

A 718-kb DNA Sequence of the Escherichia coli K-12 Genome Corresponding to the 12.7-28.0 min Region on the Linkage Map

The 718,122 base pair (bp) sequence of the Escherichia coliK-12 genome corresponding to the region from 12.7 to 28.0 minuteson the genetic map is described. This region contains at least682 potential open reading frames, of which 278 (41%) have beenpreviously identified, 147 (22%) were homologous to other knowngenes, 138 (20%) are identical or similar to the hypotheticalgenes registered in databases, and the remaining 119 (17%) didnot show a significant similarity to any other gene. In thisregion, we assigned a cluster of cit genes encoding multienzymecitrate lyase, two clusters of fimbrial genes and a set of lysogenicphage genes encoding integrase, excisionase and repressor inthe e14 genetic element. In addition, a new valine tRNA gene,designated valZ, and a family of long directly repeated sequences,LDR-A, -B and -C, were found.     

A Role for Arabidopsis PUCHI in Floral Meristem Identity and Bract Suppression

At the onset of flowering, the Arabidopsis thaliana primary inflorescence meristem starts to produce flower meristems on its flank. Determination of floral fate is associated with changes in the growth pattern and expression of meristem identity genes and suppression of a subtending leaf called a bract. Here, we show a role in floral fate determination and bract suppression for the PUCHI gene, an AP2/EREBP family gene that has previously been reported to play roles in lateral root morphogenesis. Mutations in PUCHI cause partial conversion of flowers to inflorescences, indicating that PUCHI is required for flower meristem identity. PUCHI is transiently expressed in the early flower meristem and accelerates meristem bulging while it prevents the growth of the bract primordium. The function of PUCHI in floral fate determination and bract suppression overlaps that of the BLADE-ON-PETIOLE1 (BOP1) and BOP2 genes, which encode a pair of redundant regulatory proteins involved in various developmental processes, including leaf morphogenesis and flower patterning. We also show that PUCHI acts together with BOP1 and BOP2 to promote expression of LEAFY and APETALA1, two central regulators of floral meristem identity. Expression patterns of the PUCHI and BOP genes point to a role in spatial control of flower-specific activation of these meristem identity genes.     

Principal component and hierarchical clustering analysis of metabolites in destructive weeds; polygonaceous plants

Comprehensive analysis of metabolites using capillary electrophoresis–mass spectrometry was carried out in harmful weeds belonging to Polygonaceae. A principal component analysis revealed clear distinctions among eight Rumex species and Fallopia japonica. Hierarchical clustering data showed that respective metabolites can be grouped due to species differences. There was a positive relationship between oxalate and citrate, oxalate and ascorbate, and oxalate and glutamine. The amount of oxalate per leaf fresh weight was not affected by increased concentrations of exogenously supplied nutrients from Hoagland’s formulation in one of the most destructive weeds R. obtusifolius. The oxalate accumulation in this plant is independent of external nutrient level, where nutrient-rich environments apparently stimulate internal constituents such as amino acids and other metabolites.     

Supplementation of endothelial cells with mitochondria-targeted antioxidants inhibit peroxide-induced mitochondrial iron uptake, oxidative damage, and apoptosis

The mitochondria-targeted drugs mitoquinone (Mito-Q) and mitovitamin E (MitoVit-E) are a new class of antioxidants containing the triphenylphosphonium cation moiety that facilitates drug accumulation in mitochondria. In this study, Mito-Q (ubiquinone attached to a triphenylphosphonium cation) and MitoVit-E (vitamin E attached to a triphenylphosphonium cation) were used. The aim of this study was to test the hypothesis that mitochondria-targeted antioxidants inhibit peroxide-induced oxidative stress and apoptosis in bovine aortic endothelial cells (BAEC) through enhanced scavenging of mitochondrial reactive oxygen species, thereby blocking reactive oxygen species-induced transferrin receptor (TfR)-mediated iron uptake into mitochondria. Glucose/glucose oxidase-induced oxidative stress in BAECs was monitored by oxidation of dichlorodihydrofluorescein that was catalyzed by both intracellular H(2)O(2) and transferrin iron transported into cells. Pretreatment of BAECs with Mito-Q (1 microM) and MitoVit-E (1 microM) but not untargeted antioxidants (e.g. vitamin E) significantly abrogated H(2)O(2)- and lipid peroxide-induced 2',7'-dichlorofluorescein fluorescence and protein oxidation. Mitochondria-targeted antioxidants inhibit cytochrome c release, caspase-3 activation, and DNA fragmentation. Mito-Q and MitoVit-E inhibited H(2)O(2)- and lipid peroxide-induced inactivation of complex I and aconitase, TfR overexpression, and mitochondrial uptake of (55)Fe, while restoring the mitochondrial membrane potential and proteasomal activity. We conclude that Mito-Q or MitoVit-E supplementation of endothelial cells mitigates peroxide-mediated oxidant stress and maintains proteasomal function, resulting in the overall inhibition of TfR-dependent iron uptake and apoptosis.     

International Commission for Protection against Environmental Mutagens and Carcinogens. ICPEMC working paper 5/1: Perspectives in mutation epidemiology. 1. Incidence and prevalence of genetic disease (excluding chromosomal aberrations) in human populations

Detailed knowledge of the birth frequency or the cumulative incidence over all ages of genetic diseases in human populations is a prerequisite for assessing the magnitude of possible genetic hazards caused by environmental mutagens. However, both theoretical and practical difficulties are involved in precisely measuring the total frequency of these diseases. Two sets of data from large-scale population studies, one from Northern Ireland and the other from British Columbia, are compared with each other and with the results from ad hoc surveys for individual monogenic disorders. With due allowance for differences in approach, examination indicates that the data from the large-scale population studies are inadequate. However, it could provide a crude estimate of the total frequency of genetic diseases and a fairly reliable estimate of the individual frequency of certain genetic disorders with early onset that are familiar and readily diagnosed. In addition to environmental mutagens, there are a number of factors associated with current human activity that may change the incidence of genetic diseases. In order to monitor the human population for environmental mutagens, the change in frequency of sporadic cases of those genetic diseases that arose from fresh mutation and that can be easily detected as early as possible should be followed closely. The mechanism of data collection currently being employed in some countries for childhood cancers, certain congenital malformations, and inborn errors of metabolism could be extended to include the so-called sentinel phenotypes. The rationale and feasibility of using retinoblastoma and Wilms' tumor (nephroblastoma) as examples of such population monitoring are described.     

Fas-mediated apoptosome formation is dependent on reactive oxygen species derived from mitochondrial permeability transition in Jurkat cells

Generation of reactive oxygen species (ROS) and activation of caspase cascade are both indispensable in Fas-mediated apoptotic signaling. Although ROS was presumed to affect the activity of the caspase cascade on the basis of findings that antioxidants inhibited the activation of caspases and that the stimulation of ROS by itself activated caspases, the mechanism by which these cellular events are integrated in Fas signaling is presently unclear. In this study, using human T cell leukemia Jurkat cells as well as an in vitro reconstitution system, we demonstrate that ROS are required for the formation of apoptosome. We first showed that ROS derived from mitochondrial permeability transition positively regulated the apoptotic events downstream of mitochondrial permeability transition. Then, we revealed that apoptosome formation in Fas-stimulated Jurkat cells was clearly inhibited by N-acetyl-L-cysteine and manganese superoxide dismutase by using both the immunoprecipitation and size-exclusion chromatography methods. To confirm these in vivo findings, we next used an in vitro reconstitution system in which in vitro-translated apoptotic protease-activating factor 1 (Apaf-1), procaspase-9, and cytochrome c purified from human placenta were activated by dATP to form apoptosome; the formation of apoptosome was markedly inhibited by reducing reagents such as DTT or reduced glutathione (GSH), whereas hydrogen peroxide prevented this inhibition. We also found that apoptosome formation was substantially impaired by GSH-pretreated Apaf-1, but not GSH-pretreated procaspase-9 or GSH-pretreated cytochrome c. Collectively, these results suggest that ROS plays an essential role in apoptosome formation by oxidizing Apaf-1 and the subsequent activation of caspase-9 and -3.